Tuesday, September 13, 2011

Glowing Green Cells

Just before the weekend, my professor wanted to look under the microscope to see whether our transformed diatom cells were expressing our reporter gene GFP. I've been culturing several different lines of transformed diatoms since the transformation. These cells were co-transformed, which means the resistance plasmid is separate from the GFP plasmid--that is the transformation process shot two different plasmids at the diatom cultures. Therefore we need to select for diatom lines that posses both plasmids: the resistance plasmid and the GFP plasmid.

Below is a poster I made briefly describing each of the three main steps in the process: transformation, growing the diatoms on selective agar plates, and subculturing diatom cells in selective liquid cultures.

The diatoms will only grow if they received a copy of the resistance plasmid. And well, we can tell that the diatoms don't particularly like the antibiotic we put in the growth media:

By performing a simple antibiotic sensitivity test, we can show that diatoms will not survive in the presence of the antibiotic unless they receive the plasmid that confers resistance.

Below is the first round of pictures of cells expressing GFP. These pictures came from a single colony on an agar plate, which means I didn't transfer it to a liquid culture (i.e. it was sacrificed for science). However it does give us promise that we will have GFP-expressing lines of diatoms in the near future/ I've been working on growing up the cells in liquid cultures and hope to have something by the end of the week. After that, it's a matter of growing up larger liquid cultures before we can test some science!

The view through the microscope. So much green!

This frame shows some of the variability in the intensities of fluorescence. Denser cell populations will appear brighter.

Some clumps of cells are larger than others.

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